The goal of this project is to understand and predict the structure and the conformational flexibility of RNA molecules as a function of base sequence and solution conditions. To this end, ribo-oligonucleotides containing the various structural features found in naturally-occurring nucleic acids will be synthesized and investigated by microcalorimetric techniques. Flow and batch calorimetry will be used to isothermally study conformational transitions induced by salt, pH and divalent metal ions. Transition calorimetry will be used to investigate thermally-induced conformational changes. Particular attention will be directed towards determining the effect of modified bases on the stability of single- and double-stranded structures. These calorimetric experiments will supply a complete, model-independent thermodynamic profile of helical and nonbonded regions in RNA molecules as a function of base sequence (including modified bases), salt, pH, divalent metal ions and temperature. These data will provide an empirical basis for evaluating structure stability and for predicting conformational flexibility. These conformational experiments will also provide us with fundamental insights into the nature of order-disorder transitions in RNA molecules. A comparison of the calorimetric and the van't Hoff enthalpy values will enable us to assess the applicability of the two-state model to the transitions being studied. For transitions in which intermediate states are significantly populated, we will evaluate the possibility of terminal base pair fraying ("end-effects") and use the calorimetric data to calculate the size of the cooperative unit.